Abstract

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Purpose: We developed and validated a liquid chromatography/mass spectrometric (LC/MS) method and performed pharmacokinetic studies in mice, rats, and monkeys to define the plasma pharmacokinetics and bioavailability of zebularine, an inhibitor of DNA methyltransferase being considered for clinical evaluation. Experimental Design: Stability and protein binding of zebularine were characterized in mouse, rat, dog, monkey, and human plasma. Pharmacokinetic studies were performed in CD2F1 mice (100 mg/kg i.v. or 1,000 mg/kg p.o.), Fischer 344 rats (50 mg/kg i.v., 250 mg/kg p.o., or 500 mg/kg p.o.), and rhesus monkeys (either 500 mg/kg or 1,000 mg/kg i.v. and p.o.). Plasma zebularine concentration versus time data were analyzed with non-compartmental and compartmental methods. Interspecies scaling was performed according to standard methods. Results: In mice, plasma zebularine concentrations declined with terminal half-lives (t1/2) of 40 min and 91 min after 100 mg/kg i.v. and 1,000 mg/kg p.o., respectively. Zebularine plasma concentration versus time curves (AUC) after 100 mg/kg i.v. and 1,000 mg/kg p.o.doses were 7,323 μg/ml · min and 4,935 μg/ml · min, respectively, corresponding to a total body clearance (Cltb) of 13.65 ml/min/kg, an apparent total body clearance (Clapp) of 203 ml/min/kg, and an oral bioavailability of 6.7%. In rats, plasma zebularine concentration declined with t1/2 of 363 min, 110 min, and 126 min after 50 mg/kg i.v., 250 mg/kg p.o., and 500 mg/kg p.o. doses, respectively. The zebularine AUC’s after 50 mg/kg i.v., 250 mg/kg p.o., and 500 mg/kg p.o. doses were 12,526 μg/ml · min, 1,969 μg/ml · min, and 7,612 μg/ml · min, corresponding to a Cltb 3.99 ml/min/kg for the 50 mg/kg i.v. dose and Clapp of 127 and 66 ml/min/kg for the 250 mg/kg and 500 mg/kg p.o. doses, respectively. Oral bioavailabilities of 3.1% and 6.1% were calculated for the 250 mg/kg and 500 mg/kg p.o. doses, respectively. In rhesus monkeys, the zebularine t1/2 ranged from 70 to 150 min, Cltb ranged from 3.55 to 10.85 ml/min/kg after i.v. administration and Clapp ranged from 886 to 39,572 ml/min/kg after oral administration of 500 and 1,000 mg/kg. Zebularine oral bioavailability was less than 1% in each of the four monkeys studied. Species scaling produced a relationship described by the equation: CLtb = 6.46 (weight0.9) Conclusions: Zebularine has limited oral bioavailability, which is lower in non-human primates than in rodents. Interspecies scaling projects a zebularine CLtb of 296 ml/min in a 70-kg human, but clinical studies will be required to characterize the actual CLtb and oral bioavailability in humans. The LC/MS assay developed should be applicable in those studies.